CN106674329B - Peptides having antimicrobial, anticancer/wound healing promoting activity and uses thereof - Google Patents
Peptides having antimicrobial, anticancer/wound healing promoting activity and uses thereof Download PDFInfo
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Abstract
The present invention provides a peptide having antimicrobial, anticancer and/or wound healing promoting activity, comprising: an alpha-helical (alpha-helix) peptide; and a short peptide consisting of about 4 to 10 positively charged amino acids linked to the N-terminus of the alpha-helical peptide to form the peptide having antimicrobial, anticancer and/or wound healing activity, wherein the total length of the peptide having antimicrobial, anticancer and/or wound healing activity is about 10 to 20 amino acids.
Description
Technical Field
The present invention relates to a novel peptide, and more particularly to a peptide having antimicrobial, anticancer and/or wound healing promoting activities, a pharmaceutical composition comprising the same, and uses of the same.
Background
Cationic Antimicrobial peptides are important for the regulation of the innate immune system of plants, insects and animals (Zasloff M (2002) Antimicrobial peptides of multicell organisms. Nature 415:389-395.), and were originally recognized as candidates for antibacterial and antifungal activity (Rothstem DM, Spacipoli P, Tran LT, Xu T, Roberts FD, et al (2001) Antimicrobial activity recovery in P-113, a 12-amino-acid acquisition of histatin 5. Antimicrobial activities Chemother45: 1367-1373; Yu-. Cationic Antimicrobial peptides are generally characterized by their positive charge and amphoteric properties, which enable them to bind to negatively charged bacterial cell membranes, leading to cell membrane rupture and thus bacterial death (Hancock RE and Sahl HG (2006) Antimicrobial and host-defective peptides as new anti-infectious therapeutic peptides. nat Biotechnol 24: 1551. 1557. La Rocca P, Shai Y and Sansom MS (1999) Peptide-bilayer interactions: ligands of dermaseptin B, an Antimicrobial Peptide. biophysis Chem 76: 145. 159.). The cell membrane lytic properties of cationic antimicrobial peptides make them potential therapeutic agents for overcoming the problem of antibiotic resistance (Yu HY, Huang KC, Yip BS, Tu CH, Chen HL, et al (2010) Rational design of tryptophan-rich antimicrobial peptides with enhanced antimicrobial activity and specificity. ChemB chembiochem 11: 2273-.
Although great effort has been devoted to the development of new treatment modalities, Cancer currently remains the leading cause of death (Siegel R, Ma J, Zou Z and Jemal a (2014) Cancer standards 2014.CA Cancer J Clin 64: 9-29). However, although chemotherapy has side effects on normal cells and tissues and is prone to develop multidrug resistance, they remain the primary drugs for treating Cancer in the later stages and metastatic stages (Siegel R, Ma J, Zou Z and Jemal a (2014) Cancer standards 2014.CA Cancer J Clin 64: 9-29).
Thus, novel cancer drugs with low toxicity to normal cells and a novel model of mechanism that avoids multidrug resistance may provide a new direction for anticancer therapy.
Therefore, there is a need for a novel peptide drug with low toxicity to normal cells and with antimicrobial and anticancer activity.
Disclosure of Invention
The present invention provides a peptide having antimicrobial, anticancer and/or wound healing promoting activity, comprising: an alpha-helical (alpha-helix) peptide; and a short peptide consisting of about 4 to 10 positively charged amino acids linked to the N-terminus of the alpha-helical peptide to form the peptide having antimicrobial, anticancer and/or wound healing activity, wherein the total length of the peptide having antimicrobial, anticancer and/or wound healing activity is about 10 to 20 amino acids.
The present invention also provides a pharmaceutical composition comprising: the aforementioned peptides having antimicrobial, anticancer and/or wound healing promoting activity; and a pharmaceutically acceptable carrier or salt, wherein the pharmaceutical composition has antimicrobial activity, has anticancer activity and/or has wound healing promoting activity, and does not affect normal cells.
The invention also provides the use of the peptide with antimicrobial, anticancer and/or wound healing promoting activity for preparing an antimicrobial, anticancer and/or wound healing promoting drug, wherein the drug has antimicrobial activity, anticancer activity and/or wound healing promoting activity, and does not affect normal cells.
In order to make the aforementioned and other objects, features and advantages of the present invention comprehensible, preferred embodiments accompanied with figures are described in detail as follows:
drawings
FIG. 1 shows the color grading (shown in grey scale) of Ampicillin (AP) and the Minimum Inhibitory Concentrations (MIC) of the S1, Nal2-S1, K4R2-Nal2-S1 and K6-Nal2-S1 peptides against E.coli (Escherichia coli), Staphylococcus aureus (Staphylococcus aureus) and Pseudomonas aeruginosa (Pseudomonas aeruginosa) at different NaCl concentrations.
FIG. 2A shows the survival of PC9 and PC9-G cells after 3 hours of treatment with S1, Nal2-S1, K4R2-Nal2-S1 or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1 or K6-Nal2-S1 against the activity of PC9 and PC9-G cell strains).
FIG. 2B shows the survival of A549 and OECM-1 cells after 3 hours of treatment with the S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 against the activity of A549 and OECM-1 cell lines).
FIG. 2C shows the survival of SAS and C9 cells after 3 hours of treatment with the S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 against the activity of SAS and C9 cell strains).
FIG. 3A shows the survival of PC9 and PC9-G cells after 12 hours of treatment with S1, Nal2-S1, K4R2-Nal2-S1 or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1 or K6-Nal2-S1 against the activity of PC9 and PC9-G cell strains).
FIG. 3B shows the survival of A549 and OECM-1 cells after 12 hours of treatment with the S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 against the activity of A549 and OECM-1 cell lines).
FIG. 3C shows the survival of SAS and C9 cells after 12 hours of treatment with S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 against the activity of SAS and C9 cell strains).
FIG. 4A shows the survival of PC9 and PC9-G cells after 24 hours of treatment with S1, Nal2-S1, K4R2-Nal2-S1 or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1 or K6-Nal2-S1 against the activity of PC9 and PC9-G cell strains).
FIG. 4B shows the survival of A549 and OECM-1 cells after 24 hours of treatment with the S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 against the activity of A549 and OECM-1 cell lines).
FIG. 4C shows the survival of SAS and C9 cells after 24 hours of treatment with S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 peptides (S1, Nal2-S1, K4R2-Nal2-S1, or K6-Nal2-S1 against the activity of SAS and C9 cell strains).
FIG. 5A shows the hemolytic properties of the S1, Nal2-S1, K4R2-Nal2-S1 and K6-Nal2-S1 peptides on human erythrocytes.
FIG. 5B shows the cytotoxicity of S1, Nal2-S1, K4R2-Nal2-S1 and K6-Nal2-S1 peptides against Human Fibroblast (HFW).
FIG. 6A shows representative phase contrast images of PC9 and HFW cells treated with or without 12 μ M FITC-K4R2-Nal2-S1 for 5 minutes.
FIG. 6B shows the results of immunofluorescence analysis of the interaction of FITC-conjugated-K4R 2-Nal2-S1 peptide with PC9 cells and HFW cells. For nuclear detection, cells were pre-stained with DAPI, followed by treatment with FITC-K4R2-Nal2-S1 peptide (12 μ M) for 1 hour. DAPI and FITC exhibit blue and green signals under UV and blue light sources, respectively. All scales are 20 μm.
FIG. 7 shows Western blot analysis of activated caspase 3 expression to monitor apoptosis (apoptosis) in PC9 cells and HFW cells at the indicated time points. GAPDH was used as a loaded control.
Fig. 8A shows that male nude mice injected subcutaneously with PC9 human lung cancer cells and intravenously with K4R2Nal2-S1 peptide (right panel) or PBS control (left panel) were dorsal at day 46 after cancer cell transplantation (tumor growth was allowed at day 5, then treated for day 40, and photographed at day 46).
Figures 8B and 8C show body weight (figure 8B) and tumor volume (figure 8C) of the nude mice in figure 8A during the indicated monitoring period.
Figure 8D shows the exposed tumors of mice treated with K4R2Nal2-S1 peptide (top) or PBS (bottom) (mice were sacrificed at day 46 post cancer cell implantation). 12 exposed tumors were found in the PBS treated group (6 mice implanted x2 side), but only 7 exposed tumors were found in the K4R2Nal2-S1 peptide treated group.
FIG. 8E shows the total weight of tumors in the K4R2Nal2-S1 peptide-treated mice of FIG. 8D relative to the total weight of tumors in PBS-treated mice.
Figure 9 shows that xenograft tumors excised from the mice of figures 5A and 5B were hematoxylin-eosin stained (H & E stain) with immunohistochemistry staining (IHC) for expression of cleaved PARP to monitor apoptosis. Scale bars 500 μm, 100 μm and 50 μm (40X, 200X and 400X), respectively.
FIG. 10A shows images of different test groups observed at 0, 12 and 24 hours in a cell migration assay.
FIG. 10B shows the repair rates of different test groups in the cell migration analysis.
Detailed Description
In one aspect of the invention, the invention provides novel peptides having antimicrobial, anticancer and/or wound healing promoting activity.
The peptides of the present invention having antimicrobial, anticancer and/or wound healing activities may include, but are not limited to, an alpha-helix (alpha-helix) peptide and a short peptide, wherein the short peptide may be composed of about 4 to 10 positively charged amino acids and is linked to the N-terminus of the alpha-helix peptide to form the peptide of the present invention.
The total length of the peptide of the present invention may be about 10 to 20 amino acids, for example, 10, 11, 12, 13, 14, 15, 16, 17, 18 or 19 amino acids, but is not limited thereto.
The amino acids constituting the α -helical peptide are not particularly limited, and may be all natural amino acids or all unnatural amino acids, or may contain both natural amino acids and unnatural amino acids, as long as the peptide composed of these amino acids can form an α -helical structure. In other words, the sequence of the above-mentioned α -helical peptide is not particularly limited as long as the amino acids constituting the sequence can form an α -helical structure.
In one embodiment, the α -helical peptide may have one or more unnatural amino acids, but is not limited thereto. Each of the one or more unnatural amino acids may be independently, for example, naphthylalanine (β -naphthylalanine, Nal), (3-benzothienyl) -alanine ((benzothiazen-3-yl) alanine, Bal), diphenylalanine (Dip), 4 '-biphenylalanine (4,4' -bipheny-yl) alanine, Bip), (9-anthryl) -alanine (anthracen-9-yl) alanine, Ath), or (2,5,7-tri-tert-butyl-indol-3-yl) alanine ((2,5,7-tri-tert-butyl-indol-3-yl) alanine) (Tht), but is not limited thereto. In one embodiment, the one or more unnatural amino acids described above can be naphthylalanine.
Further, one or more unnatural amino acids in the α -helical peptide may be completely or partially arranged in series. As used herein, the term "fully contiguous" means that there are no natural amino acids between any two unnatural amino acids in the above-described alpha-helical peptide. The term "partially continuous sequence" means that, in the α -helical peptide, any two unnatural amino acids do not have any natural amino acid between them for a part of the unnatural amino acids, and the amino acids directly linked to the respective unnatural amino acids for the other part of the unnatural amino acids are natural amino acids.
Alternatively, more than one unnatural amino acid in the α -helical peptide can be arranged non-consecutively. The term "non-continuous arrangement" means that, in the above-mentioned α -helical peptide, the amino acid directly linked to each unnatural amino acid is necessarily a natural amino acid.
In one embodiment, the one or more unnatural amino acids are in a perfect sequence. In this embodiment, the one or more unnatural amino acids can be located at the N-terminus of the alpha-helical peptide and directly linked to the short peptide, or can be located at the C-terminus of the alpha-helical peptide, or can be located in the middle region of the alpha-helical peptide.
In a specific embodiment, the one or more unnatural amino acids are in a perfect sequence and are located at the N-terminus of the α -helical peptide and directly linked to the short peptide. In this particular embodiment, the α -helical peptide has two unnatural amino acids, and both unnatural amino acids can be naphthylalanine.
In another particular embodiment, the sequence of the alpha-helical peptide may comprise the sequence identification numbers: 1, for example, the sequence of an α -helical peptide can be represented by the sequence identifier: 1, but is not limited thereto.
In the peptide having antimicrobial, anticancer and/or wound healing promoting activity of the present invention, the positively charged amino acids constituting the short peptide are not particularly limited as long as they are positively charged amino acids. Each positively charged amino acid of the short peptide can be, for example, lysine, arginine, histidine, or the like, but is not limited thereto.
In one embodiment, the length of the short peptide in the peptide having antimicrobial, anticancer and/or wound healing promoting activity of the present invention may be six amino acids. In this embodiment, the six positively charged amino acids comprising the above-described short peptide may all be lysine. Alternatively, in this embodiment, the six positively charged amino acids comprising the above-described short peptide may be a combination of 4 lysines and 2 arginines. Also, in a specific embodiment, the six positively charged amino acids constituting the short peptide are a combination of 4 lysines and 2 arginines, and from the N-terminus to the C-terminus of the short peptide, these positively charged amino acids are 4 lysines and 2 arginines in order.
In addition, the N-terminal amino acid of the peptide of the present invention having antimicrobial, anticancer and/or wound healing promoting activities may be acetylated. Alternatively, the C-terminal amino acid of the peptide of the present invention having antimicrobial, anticancer and/or wound healing promoting activity may be amidated. Alternatively, the N-terminal amino acid and the C-terminal amino acid of the peptide of the present invention having antimicrobial, anticancer and/or wound healing promoting activities may be acetylated and amidated, respectively.
In one embodiment, the sequence of the peptide of the present invention having antimicrobial, anticancer and/or wound healing promoting activity may include the sequence identifiers: 2 or sequence identifier: 3, but is not limited thereto. In a particular embodiment, the sequence of the peptide of the invention having antimicrobial, anticancer and/or wound healing promoting activity is represented by seq id no: 2, or a sequence of the same. In yet another specific embodiment, the sequence of the peptide of the present invention having antimicrobial, anticancer and/or wound healing promoting activity is represented by seq id no: 3, is a sequence of the sequence 3.
The sequence of the peptide having antimicrobial, anticancer and/or wound healing promoting activity in the present invention is represented by seq id no: 2, the sequence identifier: 2 may be further acetylated. Alternatively, in this particular embodiment, the sequence identifier: 2 may be further amidated. Yet, or, in this particular embodiment, the sequence identifier: 2, the N-terminal amino acid and the C-terminal amino acid of the sequence may be further acetylated and amidated, respectively.
Similarly, the sequence of the peptide having antimicrobial, anticancer and/or wound healing promoting activity in the present invention is represented by seq id no: 3, sequence ID: 3, or alternatively, the sequence identifier: 3 may be further amidated, and alternatively, the sequence identifier: 3, the N-terminal amino acid and the C-terminal amino acid of the sequence may be further acetylated and amidated, respectively.
The novel peptide of the present invention comprising an alpha-helical peptide and a short peptide can have the following effects, but is not limited thereto.
The above novel peptides of the present invention have antimicrobial activity. As used herein, "antimicrobial activity" refers to a peptide that alters the function or metabolism of a target microorganism, e.g., affects replication, growth, toxin production, survival, etc. In one embodiment, antimicrobial activity refers to inhibiting the growth of microorganisms. Also, in a particular embodiment, antimicrobial activity means that the peptides of the invention can poison at least one microorganism.
Examples of the microorganism described in the present invention may include, but are not limited to, bacteria, fungi, viruses, protozoa, etc., particularly, microorganisms having cells or structures of lipid bilayer membranes.
The bacteria may include, but are not limited to, gram-positive bacteria and/or gram-negative bacteria. Examples of the gram-positive bacteria include, but are not limited to, Staphylococcus aureus (Staphylococcus aureus), Staphylococcus epidermidis (Staphylococcus epidermidis), Streptococcus agalactiae (Streptococcus agalactiae), Group A Streptococcus (Group A Streptococcus), Streptococcus pyogenes (Streptococcus pyogenes), Group B gram-positive Streptococcus (Group B gram-positive Streptococcus), Listeria monocytogenes (Listeria monocytogenes), and the like. Examples of the gram-negative bacteria include, but are not limited to, Escherichia coli (Escherichia coli), Pseudomonas aeruginosa (Pseudomonas aeruginosa), Salmonella (Salmonella), Haemophilus influenzae (Haemophilus influenza), Vibrio cholerae (Vibrio cholera), Vibrio parahaemolyticus (Vibrio parahaemolyticus), Helicobacter pylori (Helicobacter pylori), and the like.
The fungi include yeasts, for example, Candida albicans (Candida albicans). The viruses may include, but are not limited to, Measles virus (Measles virus), herpes virus (HSV) (e.g., herpes virus-1 (HSV-1), herpes virus-2 (HSV-2)), Human Immunodeficiency Virus (HIV), Hepatitis C Virus (HCV), Vesicular Stomatitis Virus (VSV), visna virus (virus), Cytomegalovirus (CMV). Further, the protozoan may include Giardia (Giardia), but is not limited thereto.
Furthermore, the above novel peptides of the present invention have anticancer activity without affecting normal cells. In other words, the above novel peptides of the present invention have high selectivity for cancer cells.
As used herein, "anti-cancer activity" refers to a peptide that alters the function or metabolism of a target cancer cell, e.g., affects replication, growth, survival, etc., but is not limited thereto. In one embodiment, the anti-cancer activity is inhibition of growth of cancer cells. In a further specific embodiment, the anti-cancer activity means that the peptide of the invention can poison at least one cancer cell.
The cancer cell is not particularly limited, and may be, for example, a lung cancer cell, an oral cancer cell, a prostate cancer cell, a breast cancer cell, a liver cancer cell, a pancreatic cancer cell, or the like, but is not limited thereto. The lung cancer cell includes, but is not limited to, lung adenocarcinoma cell. Examples of the oral cancer cells include, but are not limited to, Oral Squamous Cell Carcinoma (OSCC) cells.
In addition, the above novel peptides of the present invention further have the ability to kill cancer cells resistant to anticancer drugs. In one embodiment, the above novel peptides of the invention have the ability to kill lung adenocarcinoma cells that are resistant to ereisma (gefitinib).
"wound healing" as used herein may be a continuous, dynamic, complex physiological process that may include, but is not limited to, cell proliferation, cell migration, and the like. In one embodiment, "promoting wound healing" as used herein may also mean "promoting cell proliferation" or "promoting cell migration", but is not limited thereto.
The novel peptide of the present invention can be used as an antimicrobial peptide alone, an anticancer peptide alone, a peptide for promoting wound healing alone or a peptide for simultaneously resisting microorganisms, cancers and promoting wound healing, as required, without any particular limitation.
In another embodiment of the present invention, the present invention provides a pharmaceutical composition comprising any of the aforementioned novel peptides of the present invention and a pharmaceutically acceptable carrier or salt.
The aforementioned pharmaceutical composition of the present invention has the ability to resist microorganisms, resist cancer and/or promote wound healing without affecting normal cells.
Examples of the microorganism which can be resisted by the pharmaceutical composition of the present invention may include, but are not limited to, bacteria, fungi, viruses, protozoa, etc., particularly, microorganisms having cells or structures of lipid bilayer membranes.
The bacteria may include, but are not limited to, gram-positive bacteria and/or gram-negative bacteria. The gram-positive bacteria may be, for example, but not limited to, Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus agalactiae, Streptococcus type A, Streptococcus pyogenes, Streptococcus gram-positive type B, Listeria monocytogenes, and the like. Examples of the gram-negative bacteria include, but are not limited to, Escherichia coli, Pseudomonas aeruginosa, Salmonella, Haemophilus influenzae, Vibrio cholerae, Vibrio parahaemolyticus, helicobacter pylori, etc.
The fungi include yeasts, such as Candida albicans. Such viruses may include, but are not limited to, measles virus, herpes viruses (e.g., herpes virus-1, herpes virus-2)), human immunodeficiency virus, hepatitis C virus, vesicular stomatitis virus, visna virus, cytomegalovirus. Further, the protozoa may include, but are not limited to, giardia, and the like.
Furthermore, the cancer cells may include, but are not limited to, lung cancer cells, oral cancer cells, prostate cancer, breast cancer, liver cancer, pancreatic cancer, and the like. Lung cancer cells include, but are not limited to, lung adenocarcinoma cells. Examples of oral cancer cells may include, but are not limited to, oral squamous cell carcinoma cells.
The pharmaceutical composition of the present invention further has an ability to kill cancer cells resistant to anticancer drugs. In one embodiment, the above pharmaceutical composition of the present invention has the ability to kill lung adenocarcinoma cells resistant to ereisma.
The wound healing promoting effect of the pharmaceutical composition of the present invention may also mean "promoting cell proliferation", "promoting cell migration", etc., but is not limited thereto.
In the pharmaceutical composition of the present invention, the pharmaceutically acceptable carrier can include, but is not limited to, solvents, dispersion media (dispersion medium), coating, antibacterial and antifungal agents, and isotonic and absorption delaying agents, which are compatible with pharmaceutical administration. For different modes of administration, the pharmaceutical composition can be formulated into dosage forms (dosage form) using conventional methods.
Also, the pharmaceutically acceptable salts may include, but are not limited to, salts including inorganic cations, for example, alkali metal salts such as sodium, potassium or amine salts, alkaline earth metal salts such as magnesium, calcium salts, salts containing divalent or tetravalent cations such as zinc, aluminum or zirconium salts. In addition, they may be organic salts, such as dicyclohexylamine salts and methyl-D-glucamine, amino acid salts, such as arginine, lysine, histidine, glutamine.
The drug prepared by the present invention can be administered orally, non-orally, via inhalation spray (inhalation spray), or by implantation into a reservoir (implanted reservoir). Non-oral administration may include subcutaneous (subcutaneous), intradermal (intracutaneous), intravenous (intravenous), intramuscular (intramusculary), intraarticular (intraarticular), arterial (intraspecific), intrasynovial (intrarenal), intrasternal (intrasternal), subarachnoid (intramembranous), intralesional (intradivision) injection, and perfusion techniques.
Oral compositions may be in the form of, but are not limited to, tablets, capsules, emulsions (emulsions), aqueous suspensions (aqueous suspensions), dispersions (dispersions), and solutions.
The pharmaceutical composition of the present invention can be administered to plants or animals. The above may include, but is not limited to, fish, birds, mammals, and the like. Examples of mammals can include, but are not limited to, cats, dogs, cows, horses, pigs, humans, and the like. In one embodiment, the pharmaceutical composition can be administered to a human.
In yet another embodiment of the present invention, the present invention provides the use of any of the aforementioned novel peptides of the present invention for the preparation of a medicament for antimicrobial, anticancer and/or wound healing.
Examples
A. Materials and methods
1. Statement of ethics
Human venous blood was collected from three healthy volunteers with written informed consent and with consent from the human experimental committee at the new bamboo division, Taiwan Hospital, China.
All animal experiments were performed according to the animal use guidelines of the institutional animal care and use committee (license No.: 10260) of the university of Qinghua, Taiwan. All nude mice were in CO2The animals were sacrificed and all efforts were made to minimize animal suffering.
2. Peptide preparation
Synthesizing the designed peptide. The identity (identity) of the peptide was confirmed by electrospray mass spectrometry (electrospray mass spectrometry), and the purity (> 95%) was assessed by HPLC. Peptide concentration was confirmed by using a UV/Vis spectrometer (spectrophotometer) at 280 nm. The buffer solution was prepared with distilled water of secondary glass.
3. Bacterial culture
Escherichia coli (Escherichia coli) strain (ATCC 25922), Staphylococcus aureus (Staphylococcus aureus) subsp strain (ATCC 25923, methicillin-resistant) and Pseudomonas aeruginosa (Pseudomonas aeruginosa) Migula strain (ATCC 27853, ampicillin resistance) were used to test the antibacterial activity of the peptides. The bacteria were cultured in sterile Mueller-Hinton (MH) medium at 200rpm and 37 ℃ for 8 hours.
After 8 hours of incubation, the concentration of inoculum (inoculums) was confirmed by measuring the absorption at an optical density of 600nm by UV/visible spectroscopy (OD 600 ═ 1, approximately equal to 10)8One CFU/mL)
4. Antimicrobial activity
Antibacterial activity was determined by LYM broth (broth) by the Standard broth microdilution method of the clinical laboratory standards Committee of Taiwan, China. LYM broth contains 5.4mM KCl, 5.6mM Na2HPO4,0.5mM MgSO4And 1.0mM sodium citrate (sodimucitrate). In addition, 0.4mg of ZnCl was added per liter of the medium22.0mg of FeCl3·6H2O, 0.1mg of CuSO4·5H2O, 0.1mg of MnSO4·H2O, 0.1mg of Na2B4O7·10H2O, 700mg of an amino acid mixture of leucoamino acids (tryptophan) (Clontech, Lot Number:2803C347) and 20mg of L-tryptophan. A mixture of vitamins (100X, Sigma, Product Number: M6895-100ML) and glucose were added at a final concentration of 2%.
Mu.l of peptide solution (in serial dilution ranging from 5000. mu.g/ml to 78. mu.g/ml) was prepared and mixed with 99. mu.l of inoculum (5X 10) in a polypropylene (polypropylene) 96-well plate5CFU/ml). Turbidity (turbidity) was measured at OD 600nm by an ELISA plate reader (Thermo Max, Molecular Devices, Sunnyvale, Calif.). The medium without peptide and the inoculum suspension were used as negative and positive control groups, respectively. The Minimum Inhibitory Concentration (MIC) value is the lowest concentration of peptide at which there is no significant growth (equal to or less than 90%). By using TreeView Program (Arnusch CJ, Ulm H, Josten M, Shadkchan Y, Osherov N, et al (2012) Ultrashort Peptide Bioconjugates Are obtained exclusive antibiotic Agents and synergy with Cyclodextrin and Amphotericin B. antisense Agents Chemother 56:1-9 Eisen MB, Spellman PT, Brown PO and Botstein D (1998) Cluster analysis and display of genome-wide expression patterns Proc Natl Acad Sci 95:14863-14868.) to convert the minimum inhibitory concentration to a color scale and display. All peptides were tested in triplicate.
5. Hemolytic Activity (hemolytic Activity)
Human venous blood was collected by a venous blood collection tube (BD Vacutainer, REF 367525). Serum was removed by washing with PBS buffer and centrifugation at 800g for 5 minutes. The above procedure was repeated at least three times to completely remove the serum.
Mu.l of peptide (in serial dilutions, ranging from 1.6mM to 3.1. mu.M) was mixed with 50. mu.l of 10% human red blood cells (hRBC) and incubated at 37 ℃ for 1 hour. The supernatant was collected after centrifugation at 800g for 5 minutes. The amount of hemoglobin released from human erythrocytes was confirmed by measuring the absorption at 405 nm. Negative and positive controls were represented by 10% human red blood cells not peptide-treated and 1% Triton X-100, respectively.
6. Cell culture
Human lung cancer cell strains PC9 and A549 and oral squamous cell carcinoma cell strain OECM-1 are cultured in RPMI culture medium added with 10% fetal bovine serum (total bovine serum) and antibiotics. Human tongue cancer cell line SAS, oral cancer cell line C9 and human diploid fibroblast cell line (human diploid fibroblast) HFW were cultured in DMEM medium supplemented with 10% fetal bovine serum and antibiotics. Culturing the cells in a medium containing 5% CO2In a moist incubator at 37 ℃.
A549, OECM-1, SAS, C9 and HFW cell lines were obtained from the institute of Biotechnology, Qinghua university, Taiwan, professor Yanglong. PC9 and anti-Elizabeth (gefitinib) PC9(PC9-G) cell line were obtained from the institute of Biotechnology of Qinghua university in Taiwan, China, and professor about nephrite. PC9-G was generated from culturing the PC9 cell line in Elizabeth (500nM) for 60 days.
7. Cytotoxicity
MTT assay was used to confirm cytotoxicity in vitro. All cancer cell lines were seeded at a concentration of 5000 cells/100. mu.l/well in 96-well plates and cultured for 24 hours. HFW cells were seeded at a concentration of 8000 cells/100. mu.l/well.
After removing the medium, 100 μ l of fresh medium containing peptides (ranging from 50 μ M to 3.13 μ M, HFW cells were additionally treated with 75 μ M and 100 μ M peptides) was added to the wells. After 3, 12 or 24 hours of culture, the medium was replaced with fresh medium containing MTT (0.5mg/ml) and cultured for 3 hours. After removal of medium/MTT, DMSO was added to 100 μ l to dissolve formazan (formazan) crystals. Cell viability was calculated by measuring the absorbance at 540nm using a Multi-labeled Microplate analyzer (VICTOR 3). For cancer cells, with H2O2(aq) the mixed media was positive control (cells dead), while media alone was negative control (cells alive); for normal cells, only the medium was positive control (cells were fully viable) and H was added2O2(aq) mixed media was used as negative control (cell death).
8. Image of cell survival
PC9 was mixed with HFW cells (. about.10)5Individual cells) were pre-seeded on 6-cm polystyrene (polystyrene) disks for 24 hours. The nuclei were labeled with 4',6-diamidino-2-phenylindole (4',6-diamidino-2-phenylindole, DAPI) at a final concentration of 10. mu.g/ml. After 10 min incubation, cells were washed with PBS. FITC-K4R2Nal2S1-NH at a final concentration of 12. mu.M2Add to the culture dish. After incubation at 37 ℃ for 5, 10, 20 minutes or 1 hour, cells were washed with PBS. FITC-K4R2Nal2S1-NH was visualized using an Inverted fluorescence Microscope (Inverted fluorescence Microscope Zeiss/observer. Z1)2Images in PC9 cells and HFW cells.
Western blotting (Western blotting)
PC9 cells were seeded on 10-cm polystyrene disks for 48 hours. Approximately 70% full cells were treated with 12 μ M K4R2Nal2S1 for 10 minutes, 1 hour, or 24 hours. A negative control group was prepared by treating the cells with RPMI medium without peptide for 24 hours.
Cells were harvested by mixing 200. mu.l RIPA with protease inhibitor (protease inhibitor) buffer. After centrifugation at 13000rpm for 10 minutes at 4 ℃ with ultrasonic shaking, the supernatant (cell lysate) was collected. Protein concentration of cell extracts was determined by Bradford reagent. An equal amount of boiled cell lysate (total 50. mu.g) was separated as a 10% acrylamide gel. The gel was transferred to PVDF membrane in an electro-blotting system (electro-blot system) at 300V, 350A and 80 minutes.
The membrane was incubated in blocking buffer (5% skim milk, TBST buffer) for 1 hour at room temperature and washed twice in TBST buffer. The blocked membranes were incubated with Caspase-3 antibody (EPITOMICS, clone ID: E83-77) overnight at 4 ℃, washed five times, and then incubated in secondary antibody (HRP, GeneTex catalog number: GTX21311-01) at room temperature for 1 hour. The signal is visualized by Enhanced Chemiluminescence (ECL) and recorded by a detection system (ImageQuant LAS 4000 mini).
10. Mouse and pathological study
12 male nude mice (BALB/cAnN. Cg-Foxn1nu/CrlNarl) were purchased from the laboratory animal center Taiwan (Taiwan).
Mu.l of human lung cancer cell PC9(3X106 cells) in matrigel (Corning) was injected subcutaneously into the dorsal side of 5-week-old male nude mice. Each mouse was inoculated on both sides of its back (Taguchi F, Koh Y, Koizumi F, Tamura T, Saijo N, et al (2004) Anticancer effects of ZD6474, a VEGF receptor type kinase inhibitor, in gefitinib ("Iressa") -sensitive and resistant xenograft models. cancer Sci 95: 984. sup. 989.). In implantation (cancer size)>95mm3) After 5 days, 12 mice were randomly assigned into two groups. One group received K4R2Nal2S1 peptide (5mg/Kg dissolved in 100. mu.l PBS buffer) via tail vein injection (tail vein injection) three times a week, while the other group was injected with PBS as a control group.
Body weight and cancer size were measured three times a week. Via the width2X length x 0.52 to calculate the cancer volume. The volume of cancer below 100% of the pre-treatment volume is defined as "cancer shrinkage" (Taguchi F, Koh Y, Koizumi F, Tamura T, Saijo N, et al (2004) Anticancer effects of ZD6474, a VEGF receptor tyrosine kinaseinhibitor, in gefitinib ("Iressa") -passive and reactive xenograft models, cancer Sci 95: 984-. Furthermore, the mice were dissected to confirm "cancer reduction".
After 40 days of treatment, mice were sacrificed and tumors were removed, photographed and weighed. All Animal experiments were performed according to the Animal use guidelines of the institutional Animal Care and use Committee of the university of qinghua, taiwan, and were approved by the Animal Care Committee (Animal Care Committee).
Solid cancer was fixed in 4% formaldehyde buffer solution. Paraffin-embedded tissues (paraffin-embedded tissues) were cut into 2 μm-later sections and deparaffinized in Ultraclear Buffer (J.T.Baker) and gradient ethanol (gradedethanol). The morphology of the cancer was visualized by hematoxylin-eosin stained sections (H & E stain). In addition, sections were immunostained (immunostain) with anti-cleaved-PARP (1:100) antibody (Cell Signaling, clone number: D65E 10).
Images were captured in 40X, 200X and 400X fields of view using an optical microscope (Eclipse E400, Nikon) and confocal digital microscope camera (AxioCam ic 5, ZEISS).
B. Results
1. Peptide design
Previously, the inventors have developed a strategy to increase salt tolerance and serum stability of short antimicrobial peptides by adding a naphthyl amino acid (. beta. -naphthalanine, Nal) at the end of the peptide (Chu HL, Yu HY, Yiip BS, Chih YH, Liang CW, et al (2013) Boosting saline resistance of short antimicrobial peptides, antimicrobial Agents Chemothers 57: 4050-4052.). This strategy has been successfully applied to the S1 peptide (SEQ ID NO: 4) and the ultrashort peptide KWWK (SEQ ID NO: 5).
However, peptides with naphthyl amino acid end markers have also been shown to have higher cytolytic activity (cytotoxicity). (Chu HL, Yu HY, YIp BS, Chih YH, Liang CW, et al (2013) Boosting salt resistance of short antimicrobial peptides, anti-microbial Agents Chemothers 57: 4050-. This problem can be counteracted by adding positively charged amino acids to the N-and/or C-terminus of the Antimicrobial Peptide (Yin LM, Edwards MA, Li J, Yip CM and Deber CM (2012) loops of hydrophic and Charge Distribution of Cationic Peptides in Peptide-Membrane interactions.J Biol Chem 287: 7738. 7745.).
Nal2-S1, K4R2-Nal2-S1 and K6-Nal2-S1 peptides were designed and synthesized herein and compared to their antimicrobial and anticancer activities and their cytotoxicity with the peptide S1 from which they originated. The sequences and molecular weights of the S1, Nal2S1, K4R2-Nal2-S1, and K6-Nal2-S1 peptides are listed in Table 1 below.
Table 1, primary structure of S1 peptide and analogs thereof
aNaphthyl amino acid (beta-naphthalanine, Nal)
2. Antimicrobial activity
The S1 peptide and its analogs were tested for activity against gram positive and gram negative bacteria under various salt conditions.
The minimum inhibitory concentrations of the aforementioned peptides were analyzed and the results are shown in FIG. 1. FIG. 1 shows that under LYM gravy conditions, three peptides, Nal2-S1, K4R2-Nal2-S1 and K6-Nal2-S1, were all very effective against bacteria.
Nal2-S1 and K4R2-Nal2-S1 peptides showed desirable activity under high salt conditions. However, the activity of the K6-Nal2-S1 peptide was attenuated by the addition of 100 or 200mM NaCl.
3. Cytotoxicity
The cytotoxicity of the S1 peptides and their analogs, Nal2-S1, K4R2-Nal2-S1 and K6-Nal2-S1, against human lung cancer cells (i.e., PC9, PC9-G and A549), human oral cancer cells (i.e., OECM-1, SAS and C9) and Human Fibroblast (HFW) cells at 3 hours, 12 hours and 24 hours was evaluated via MTT assay.
The 3-hour cytotoxicity of the above-mentioned peptide for various cancer cells is shown in FIGS. 2A-2C, respectively, the 12-hour cytotoxicity of the above-mentioned peptide for various cancer cells is shown in FIGS. 3A-3C, respectively, and the 24-hour cytotoxicity of the above-mentioned peptide for various cancer cells is shown in FIGS. 4A-4C, respectively.
The results showed that three peptides, Nal2-S1, K4R2-Nal2-S1 and K6-Nal2-S1, embedded with naphthyl amino acid (Nal) all had potent anticancer activity against different cancer cells after treating various cancer cells with the peptides for 24 hours (see FIGS. 4A-4C).
Similar results were also observed at earlier time points (i.e., 3 hours versus 12 hours, see FIGS. 2A-2C and 3A-3C, respectively).
The selectivity of these peptides was studied using human red blood cells (hRBCs) and Human Fibroblasts (HFW). The results are shown in FIGS. 5A and 5B.
The lytic activity of all peptides on human erythrocytes. The test was carried out at 37 ℃ in 1 hour of culture and was calculated from the minimum haemolytic concentration (minimum haemolytic concentration).
Nal2-S1 peptide showed 10% hemolytic activity at a peptide concentration of 25. mu.M. Surprisingly, the K4R2-Nal2-S1 and K6-Nal2-S1 peptides were found to be neither hemolytic nor even at a concentration of 800. mu.M. However, the cytotoxicity of these peptides on human fibroblasts was found to be the S1 peptide < K4R2-Nal2-S1 peptide < K6-Nal2-S1 peptide < Nal2-S1 peptide.
Since K4R2-Nal2-S1 peptide exhibits better salt tolerance and lower toxicity to human erythrocytes and human fibroblasts than Nal2-S1 and K6-Nal2-S1 peptides, K4R2-Nal2-S1 peptide was selected to study its anticancer activity in PC9 cancer cell line and xenograft animal model (xenograft animal model).
4. Mechanism of anti-cancer in vitro
To investigate the mode of action of the K4R2Nal2-S1 peptide on human cancer cell lines (PC9) and Human Fibroblasts (HFW), cells were treated with FITC-labeled-K4R 2-Nal 2-S1. The nuclei were labeled with DAPI and the blue signal was observed via UV excitation light. The fluorescence distribution of FITC-labeled-K4R 2-Nal2-S1 peptide on the cell membrane was visualized by inverted fluorescence microscopy.
Phase-contrast microscopy showed that the K4R2-Nal2-S1 peptide treatment induced cell expansion not in human fibroblasts, but in PC9 (see FIG. 6A). Furthermore, immunofluorescence analysis showed that treatment with FITC-labeled-K4R 2-Nal2-S1 peptide resulted in the formation of a small stain on the cell membrane in PC9 cells, but not in human fibroblasts (see FIG. 6B). Thus, in cancer cells, K4R2-Nal2-S1 binds to the cell membrane and induces cell death.
Immunoblotting indicated that treatment with the K4R2-Nal2-S1 peptide activated caspase 3 in PC9 cells but not in human fibroblasts, suggesting that K4R2-Nal2-S1 peptide-mediated cell death involves apoptosis (apoptosis) (see FIG. 7).
The above results indicate that the K4R2-Nal2-S1 peptide preferentially binds to cancer cells and causes apoptosis.
5. Inhibition of lung cancer cell growth in xenograft animal models
To evaluate the anticancer efficacy of the K4R2-Nal2-S1 peptide in vivo, PC9 cells were implanted subcutaneously into nude mice (see FIG. 8A), and then K4R2-Nal2-S1 peptide was injected into nude mice three times a week at a dose of 5mg/kg via intravenous route (see FIGS. 8B and 8C).
During administration, no weight loss was found in the K4R2Nal2-S1 peptide treated group (see 8B); however, significant inhibition in tumor growth was observed in mice treated with the K4R2-Nal2-S1 peptide alone (see FIG. 8C). Treatment with the K4R2-Nal2-S1 peptide also reduced the volume and weight of the resulting tumors 40 days after injection (see FIGS. 8D and 8E).
The above results clearly show that treatment with the K4R2-Nal2-S1 peptide attenuated the growth of xenograft tumors.
To examine the association of cellular necrosis (necrosis) and apoptosis in the K4R2Nal2-S1 peptide-mediated anticancer effect, lung tumors generated in xenograft mouse models were excised and analyzed by hematoxylin-eosin (H & E) staining and immunohistochemical staining (IHC) for cleaved PARP, a marker for apoptosis.
Hematoxylin-eosin (H & E) staining showed extensive areas of cellular necrosis in K4R2-Nal2-S1 peptide-treated cancers, but not in PBS-treated cancers (figure 9). Immunohistochemical staining indicated increased expression of cleaved PARP in the immediate necrotic region of the cells in the cancer treated with the K4R2-Nal2-S1 peptide. These results support the insight that treatment with the K4R2Nal2-S1 peptide inhibits tumor growth.
6. Cell migration (cell migration) assay (in vitro wound healing assay)
HaCaT cells (40,000) suspended in DMEM medium supplemented with 10% fetal bovine serum were seeded on each side of a culture insert (ibidi, Germany). The insert was placed in a 24-well plate. At 37 ℃ and 5% CO2After overnight incubation, the inserts were removed and 1ml serum-free DMEM supplemented with different concentrations of peptide was added. The medium containing no peptide and the medium containing 100ng/ml Epidermal Growth Factor (EGF) were a negative control group and a positive control group, respectively. Images of the damaged area (injury area) were observed at 0, 12 and 24 hours by using an inverted fluorescence microscope (Zeiss/observer. z1) (10X). Also, the wound area (wound area) was evaluated by Adobe Photoshop. Then, the repair rate (repairing rate) is calculated: repair rate (area)t0Area oft12,24) Area/areat0Where t is time.
FIG. 10A shows images of different test groups observed at 0, 12 and 24 hours in a cell migration assay. Whereas, fig. 10B shows the wound repair rate for different test groups in the cell migration analysis.
As is clear from FIGS. 10A and 10B, the K4R2-Nal2-S1 peptide at different concentrations has better wound repair ability than the negative control group. In other words, the K4R2-Nal2-S1 peptide has the activity of promoting wound healing.
Although the present invention has been described with respect to the preferred embodiments, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims.
Sequence listing
<110> Chengdu dimension
<120> peptide having antimicrobial, anticancer and/or wound healing promoting activity, pharmaceutical composition comprising the same, and use of the same
<160> 5
<170> PatentIn version 3.5
<210> 1
<211> 12
<212> PRT
<213> Artificial sequence
<220>
<223> chemically synthesized peptide
<220>
<221> MISC_FEATURE
<222> (1)..(2)
<223> Xaa is naphthylalanine (beta-naphthalanine)
<400> 1
Xaa Xaa Lys Lys Trp Arg Lys Trp Leu Ala Lys Lys
1 5 10
<210> 2
<211> 18
<212> PRT
<213> Artificial sequence
<220>
<223> chemically synthesized peptide
<220>
<221> MISC_FEATURE
<222> (7)..(8)
<223> Xaa is naphthylalanine (beta-naphthalanine)
<400> 2
Lys Lys Lys Lys Arg Arg Xaa Xaa Lys Lys Trp Arg Lys Trp Leu Ala
1 5 10 15
Lys Lys
<210> 3
<211> 18
<212> PRT
<213> Artificial sequence
<220>
<223> chemically synthesized peptide
<220>
<221> MISC_FEATURE
<222> (7)..(8)
<223> Xaa is naphthylalanine (beta-naphthalanine)
<400> 3
Lys Lys Lys Lys Lys Lys Xaa Xaa Lys Lys Trp Arg Lys Trp Leu Ala
1 5 10 15
Lys Lys
<210> 4
<211> 4
<212> PRT
<213> Artificial sequence
<220>
<223> chemically synthesized peptide
<400> 4
Lys Trp Trp Lys
1
<210> 5
<211> 10
<212> PRT
<213> Artificial sequence
<220>
<223> chemically synthesized peptide
<400> 5
Lys Lys Trp Arg Lys Trp Leu Ala Lys Lys
1 5 10
Claims (2)
1. Use of a peptide having an activity of promoting wound healing for the preparation of a medicament having an activity of promoting wound healing, wherein the medicament has an activity of promoting wound healing without affecting normal cells,
wherein the sequence of the peptide having wound healing promoting activity is seq id no: 2, or a sequence of the sequence (2).
2. The use as described in claim 1, wherein the N-terminal amino acid and the C-terminal amino acid of the peptide having the wound-healing activity are acetylated and amidated, respectively.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101622271A (en) * | 2007-01-16 | 2010-01-06 | 加利福尼亚大学董事会 | Novel antimicrobial peptides |
| CN102258767A (en) * | 2010-05-24 | 2011-11-30 | 黄敏铨 | Composition for promoting wound healing |
| CN102363040A (en) * | 2011-10-21 | 2012-02-29 | 北京锐瑟科技有限公司 | Antimicrobial peptide preparation for mucosal tissues |
| CN103097405A (en) * | 2010-06-14 | 2013-05-08 | 基因信号国际公司 | Novel peptides for wound healing |
| CN103260635A (en) * | 2010-08-23 | 2013-08-21 | 加利福尼亚大学董事会 | Compositions and uses of materials with high antimicrobial activity and low toxicity |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GR862206B (en) * | 1985-09-12 | 1986-12-31 | Univ Tulane | Therapeutic somatostatin analogs |
| US5686422A (en) * | 1987-03-17 | 1997-11-11 | Research Corporation Technologies, Inc. | Synthetic inhibitors of mammalian collagenase |
-
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- 2015-11-05 CN CN201510749184.6A patent/CN106674329B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101622271A (en) * | 2007-01-16 | 2010-01-06 | 加利福尼亚大学董事会 | Novel antimicrobial peptides |
| CN102258767A (en) * | 2010-05-24 | 2011-11-30 | 黄敏铨 | Composition for promoting wound healing |
| CN103097405A (en) * | 2010-06-14 | 2013-05-08 | 基因信号国际公司 | Novel peptides for wound healing |
| CN103260635A (en) * | 2010-08-23 | 2013-08-21 | 加利福尼亚大学董事会 | Compositions and uses of materials with high antimicrobial activity and low toxicity |
| CN102363040A (en) * | 2011-10-21 | 2012-02-29 | 北京锐瑟科技有限公司 | Antimicrobial peptide preparation for mucosal tissues |
Non-Patent Citations (3)
| Title |
|---|
| Boosting Salt Resistance of Short Antimicrobial Peptides;Hung-Lun Chu 等;《Antimicrobial Agents and Chemotherapy》;20130831;第57卷(第8期);摘要，第4050页左栏第4段、右栏第1段 * |
| Lois M. Yin 等.Roles of Hydrophobicity and Charge Distribution of Cationic Antimicrobial Peptides in Peptide-Membrane Interactions.《THE JOURNAL OF BIOLOGICAL CHEMISTRY》.2012,第287卷(第10期), * |
| Roles of Hydrophobicity and Charge Distribution of Cationic Antimicrobial Peptides in Peptide-Membrane Interactions;Lois M. Yin 等;《THE JOURNAL OF BIOLOGICAL CHEMISTRY》;20120302;第287卷(第10期);第7738页右栏第2段，第7741页右栏最后一段，第7744页左栏第1段，表1，表2 * |
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